2 [Cells contain SV40 and CMV viral DNA sequences]
Biosafety classification is based on U.S. Public Health Service Guidelines, it is the responsibility of the customer to ensure that their facilities comply with biosafety regulations for their own country.
Age
fetus
Gender
female
Applications
This cell line has been engineered to report tau seeding activity. Tau seeds introduced into the culture media of CRL-3275 can nucleate the aggregation of the endogenous tau reporter proteins. This aggregation produces a FRET signal which can be measured via microscopy, microplate readers or flow cytometry.
Images
Derivation
The Tau RD P301S FRET Biosensor cells were derived by transducing HEK293T cells with 2 separate lentivirus constructs encoding tau RD P301S-CFP and tau RD P301S-YFP. Dual-positive cells were identified by FAC sorting and were cloned and isolated using cloning cylinders.
Complete Growth Medium
The base medium for this cell line is ATCC-formulated Dulbecco's Modified Eagle's Medium, Catalog No. 30-2002. To make the complete growth medium, add the following components to the base medium:
2mM L-alanyl-L-glutamine
fetal bovine serum to a final concentration of 10%
Subculturing
Volumes used in this protocol are for 75 cm2 flasks; proportionally reduce or increase amount of dissociation medium for culture vessels of other sizes.
Remove and discard culture medium.
Add 2.0 to 3.0 ml of 0.25% (w/v) Trypsin - 0.53 mM EDTA (ATCC 30-2101) solution to flask and observe cells under an inverted microscope until cell layer is dispersed (usually within 5 to 15 minutes).
Note: To avoid clumping do not agitate the cells by hitting or shaking the flask while waiting for the cells to detach. Cells that are difficult to detach may be placed at 37°C to facilitate dispersal.
Add 6.0 to 8.0 ml of complete growth medium and aspirate cells by gently pipetting.
Add appropriate aliquots of the cell suspension to new culture vessels.